Brain imaging and electroencephalogram patterns should be reviewed, if available

ing pocket can be explored for the development of new generations of selective kinase inhibitors. Europe PMC Funders Author Manuscripts Europe PMC Funders Author Manuscripts Results SCH772984 adopts a unique kinase binding mode in ERK1/2 SCH772984 is a novel pyridine-indazole inhibitor with an unusual extended piperazinephenyl-pyrimidine decoration19. To understand the molecular mechanisms of SCH772984 selectivity we determined crystal structures of this compound with ERK1 and ERK2. Both structures were refined to high resolution, and the bound inhibitor was well defined by electron density in both structures. The binding mode of SCH772984 was conserved in ERK1/2 and revealed an intricate network of interaction across the ATP binding site. Consistent with the lack of propensity of ERK1/2 to adopt a `DFG’ out conformation, the extended linear decoration of the inhibitor did not interact with the type-II binding pocket but with a so far unseen induced binding pocked located between helix C and the phosphate binding loop . Analysis of the ERK1/2 structures suggested that tight binding of the inhibitor was due to three key interactions. First, the indazole acted as a hinge binding scaffold forming two hydrogen bonds with the hinge backbone while the pyridine nitrogen formed a hydrogen bond with lysine K114. Second, the pyrrolidine linker was positioned in proximity to the conserved active site salt bridge forming a network of direct and water mediated hydrogen bonds involving also the gatekeeper Q105 and the `DFG’ motif which adopted an `in’ conformation. Intriguingly, the P-loop tyrosine Y36 flipped into the ATP site and stacked onto the pyrrolidine ring, leading to a strong distortion of the P-loop and opening of the P-loop binding pocket. Third, the linker between the pyrrolidine and the piperazine produced a sharp kink that oriented the phenyl-pyrimidine moiety towards the P-loop pocket flanked also by the C helix. GS 4059 biological activity Interactions between the kinase and the phenyl-pyrimidine rings were limited to -stacking interaction with the C Y64 and water-mediated hydrogen bonds to the pyrimidine group. Structural comparison with the unphosphorylated, inactive20 and phosphorylated, active conformations of ERK21 demonstrated that the allosteric pocket induced by SCH772984 does not exist in either states of the kinase. The formation of the PNat Chem Biol. Author manuscript; available in PMC 2015 December 22. Chaikuad et al. Page 4 loop binding pocket in ERK1/2 involved a tilt of the PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19811788 C and significant structural distortion of the P-loop while other key structural elements such as the conserved VIAK/C salt bridge as well as the DFG motif assumed active conformations. However, the Ploop conformation with the tyrosine Y36 oriented towards the ATP site would not be compatible with ATP binding and must therefore be considered an inactive state of ERK1/2. Europe PMC Funders Author Manuscripts Europe PMC Funders Author Manuscripts SCH772984 has high selectivity for ERK1/2 Next we asked if this unique binding mode confers a high degree of selectivity. Morris et al. screened SCH772984 against a panel of kinases using enzymatic assays, identifying only few additional kinases that were inhibited with considerably weaker potency19. Here we used a comprehensive KINOMEscan panel22 to assess selectivity against 456 kinases, which confirmed high specificity of SCH772984 for ERK1/2 detecting few off-targets of significantly weaker affinity. We then performed enzymatic ass